DNA载体介导的小干扰RNA Tet-on基因表达系统的调控效应检测
发布时间：2018-11-04 13:56
【摘要】：目的观察构建的DNA载体介导的小干扰RNA Tet-on基因表达系统的调控效应。方法构建DNA载体介导的小干扰RNA(siRNA)Tet-on表达载体,pDIRS4.1 siRNA。以增强绿色荧光蛋白(EGFP)基因和白细胞介素6(IL-6)基因为目的基因,应用脂质体介导的瞬时转染方法,分别将pDIRS4.1/EGFP siRNA载体和p EGFP-N3质粒、pDIRS4.1IL-6siRNA载体和p IRESIL-6质粒共转染人成骨肉瘤细胞,加入系列浓度的四环素(Dox)后,观察其诱导效应。结果 pDIRS4.1 siRNA在人成骨肉瘤细胞中能够高效转染;随着Dox浓度升高,pDIRS4.1 siRNA载体在人成骨肉瘤细胞中对目的基因的沉默作用有较好的调控性,且目的基因的表达与Dox具有严格的剂量依赖关系,高浓度Dox时,IL-6表达水平降低约12倍。结论该研究构建的以DNA载体介导的能够高效调控siRNA沉默基因效应的Tet-On基因表达系统,能够促进其在更多模型系统中进行基因功能研究的应用。
[Abstract]:Objective to investigate the regulatory effects of small interfering RNA Tet-on gene expression system mediated by DNA vector. Methods the expression vector of small interfering RNA (siRNA) Tet-on mediated by DNA vector and pDIRS4.1 siRNA. were constructed. To enhance green fluorescent protein (EGFP) gene and interleukin 6 (IL-6) gene as target genes, pDIRS4.1/EGFP siRNA vector and p EGFP-N3 plasmid were transfected by liposome-mediated transient transfection, respectively. Human osteosarcoma cells were co-transfected with pDIRS4.1IL-6siRNA vector and p IRESIL-6 plasmid. The induction effect was observed by adding a series of tetracycline (Dox) to human osteosarcoma cells. Results pDIRS4.1 siRNA was highly transfected into human osteosarcoma cells. With the increase of Dox concentration, the silencing effect of pDIRS4.1 siRNA vector on target gene in human osteosarcoma cells was better, and the expression of target gene was in strict dose-dependent manner with Dox. The expression level of IL-6 was reduced by about 12 times. Conclusion the Tet-On gene expression system mediated by DNA vector can effectively regulate the gene effect of siRNA silencing, which can promote the application of Tet-On gene function research in more model systems.
【作者单位】： 郑州大学第一附属医院药学部;
【基金】：河南省优秀创新型科技团队医学科技攻关计划项目(No:201303013)
【分类号】：Q78
本文编号：2310022
[Abstract]:Objective to investigate the regulatory effects of small interfering RNA Tet-on gene expression system mediated by DNA vector. Methods the expression vector of small interfering RNA (siRNA) Tet-on mediated by DNA vector and pDIRS4.1 siRNA. were constructed. To enhance green fluorescent protein (EGFP) gene and interleukin 6 (IL-6) gene as target genes, pDIRS4.1/EGFP siRNA vector and p EGFP-N3 plasmid were transfected by liposome-mediated transient transfection, respectively. Human osteosarcoma cells were co-transfected with pDIRS4.1IL-6siRNA vector and p IRESIL-6 plasmid. The induction effect was observed by adding a series of tetracycline (Dox) to human osteosarcoma cells. Results pDIRS4.1 siRNA was highly transfected into human osteosarcoma cells. With the increase of Dox concentration, the silencing effect of pDIRS4.1 siRNA vector on target gene in human osteosarcoma cells was better, and the expression of target gene was in strict dose-dependent manner with Dox. The expression level of IL-6 was reduced by about 12 times. Conclusion the Tet-On gene expression system mediated by DNA vector can effectively regulate the gene effect of siRNA silencing, which can promote the application of Tet-On gene function research in more model systems.
【作者单位】： 郑州大学第一附属医院药学部;
【基金】：河南省优秀创新型科技团队医学科技攻关计划项目(No:201303013)
【分类号】：Q78
【相似文献】
相关期刊论文 前2条
1 冉宇欣;伤寒杆菌免疫原制剂和DNA载体用作防御伤寒杆菌、霍乱弧菌和大肠杆菌的菌苗[J];国外医学.预防.诊断.治疗用生物制品分册;1986年01期
2 ;[J];;年期
，本文编号：2310022
本文链接：https://www.wllwen.com/kejilunwen/jiyingongcheng/2310022.html
最近更新
教材专著
